用长链RT-PCR技术分析一个完全型雄激素不敏感综合征家系AR基因的突变

Analysis of AR gene mutation in a family affected with complete androgen insensitivity syndrome using long chain RT-PCR

目的分析一例完全型雄激素不敏感综合征（complete androgen insensitivity syndrome，CAIS）患者及其家系成员雄激素受体（androgen receptor，AR）基因的潜在突变。方法提取患者及其家系成员的外周血基因组DNA和总RNA，应用PCR技术扩增X染色体上AR基因的7个外显子，对产物直接进行DNA测序，寻找突变位点；应用PCR-限制性片段长度多态性方法分析相应的基因组片段，进一步确证AR基因的突变位点；用逆转录PCR扩增AR基因，再次验证其突变位点。结果发现患者及其舅舅AR基因第7外显子的第2880个核苷酸G被A替换，引起错义突变，导致第840位密码子CGT突变为CAT，即精氨酸被组氨酸替代。该突变可能引起雄激素受体结合能力的变化，导致CAIS。患者母亲一条AR等位基因的第840位密码子亦存在CGT→CAT改变，另一条等位基因则未发现突变。结论AR基因第7外显子核苷酸序列2880位点碱基G引起的错义突变很可能是引起CAIS的原因。应用长链RT-PCR技术扩增AR基因可为CAIS的分子诊断提供新的方法。

Objective To identify potential mutation of androgen receptor （AR） gene in a patient with complete androgen insensitivity syndrome （CAIS） and his family members. Methods Total RNA and genomic DNA were extracted from the peripheral blood samples derived from the proband and her family members. Sequences of 7 exons of the AR gene were amplified with reverse transcriptase PCR（RT-PCR） and subjected to direct sequencing. Suspected mutation was also analyzed with PCR-restriction fragment length polymorphism （PCR-RFLP） and direct sequencing. Results DNA sequencing has revealed a nucleotide change （2880A;G） in the pedigree, which resulted in a missense mutation （R840H）. Conclusion A prenatal diagnostic method was established for detecting mutation of the AR gene in the pedigree. Long chain RT-PCR was first used for the detection of AR gene mutations.